Selenium is an essential trace element that has roles in cancer prevention, the immune system, brain function, and male reproduction. This element is present in proteins in the form of selenocysteine residue, which is co-translationally inserted into proteins in response to UGA codons. Mutations or single nucleotide polymorphisms in several selenoprotein genes and selenocysteine machinery genes have been associated with disease. Selenocysteine insertion is dependent on the SECIS element, a stem-loop structure present in 3'-untranslated regions of selenoprotein genes. Because UGA codon normally functions as a stop signal, selenoprotein genes are often missed by genome annotation programs. We developed bioinformatics tools that allow identification of selenoprotein genes by searching for SECIS elements and selenocysteine/cysteine pairs in homologous sequences. Using these tools, we identified sets of human and mouse selenoproteins (selenoproteomes). We suggest that in order to understand biological and biomedical effects of dietary selenium, information on identifies and functions of all selenoproteins is required. We propose to address critical questions in the area of selenium biology and its role in human health, including dependence of selenocysteine insertion on the location of UGA codon within coding sequences of selenoprotein genes, comprehensive characterization of mammalian selenoproteomes through bioinformatics and imaging approaches, and analysis of biological functions of an emerging class of endoplasmic reticulum-resident selenoproteins. The three specific aims of this application are as follows: (1) Position-dependent insertion of selenocysteine into mammalian selenoproteins. (2) Characterization of mammalian selenoproteomes. (3) Roles of Selenoprotein S and Selenoprotein K in mammalian endoplasmic reticulum-associated protein degradation. A combination of cell biology, biochemistry, bioinformatics, imaging and animal model approaches will be used to address these questions. PUBLIC HEALTH RELEVANCE: In mammals, selenium is an essential trace element due to its presence in proteins in the form of the 21st amino acid, selenocysteine. Selenium deficiency or mutations in selenoprotein or selenocysteine machinery genes may lead to disease. Our previous studies led to the identification of a set of selenoprotein genes in humans and mice. We now propose studies to understand regulation of selenocysteine insertion into proteins, characterize the mammalian selenoproteome, and determine functions of a widely occurring class of mammalian endoplasmic reticulum selenoproteins.